Nuclear IRS-1-DNA repair and mutagenesis in medulloblastoma

髓母细胞瘤中的核 IRS-1-DNA 修复和诱变

• 批准号: 7799179
• 负责人: Krzysztof Reiss
• 金额: $ 26.03万
• 依托单位: LSU HEALTH SCIENCES CENTER
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-03-06 至 2013-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7799179
• 关键词: Address; Affect; Attenuated; Benign; Binding; Binding Sites; Biological Assay; Biopsy; Cell Fractionation; Cell Nucleus; Cells; Central Nervous System Neoplasms; Cerebellar Neoplasms; Childhood; Childhood Cerebellar Neoplasm; Clinical; Collaborations; Collection; Complex; Computer software; Cytoplasmic Granules; DNA Damage; DNA Repair; DNA Repair Inhibition; DNA repair protein; Defect; Development; Epigenetic Process; Estradiol; Estrogen Receptor beta; Estrogen Receptors; Evaluation; Event; Figs - dietary; Frozen Sections; Future; Gene Expression; Genetic; Genomic Instability; Green Fluorescent Proteins; Growth; Human; Hyperplasia; ICI 182780; In Vitro; Insulin-Like Growth Factor I; Insulin-Like Growth Factor Receptor; Insulin-Like-Growth Factor I Receptor; JC Virus; Laboratories; Large T Antigen; Letters; Malignant - descriptor; Malignant Neoplasms; Malignant neoplasm of cerebellum; Mediating; Methodology; Microscopy; Molecular; Molecular Target; Mutagenesis; Mutation; Neuraxis; Nuclear; Nuclear Translocation; Oncogenic; Paraffin Embedding; Pathway interactions; Patients; Polyomavirus; Proteins; Receptor Signaling; Recombinant IGF-I; Recurrence; Reporter; Research; Research Personnel; Research Proposals; Sampling; Series; Signal Pathway; Signal Transduction; Signaling Molecule; Site; Source; System; Testing; Therapeutic; Thick; Tissue Microarray; Transgenic Mice; Tumor Tissue; Universities; Viral Tumor Antigens; base; cancer cell; clinically relevant; homologous recombination; inhibitor/antagonist; insulin receptor substrate 1 protein; medulloblastoma; medulloblastoma cell line; mutant; neoplastic cell; novel therapeutics; outcome forecast; overexpression; prevent; protein distribution; protein protein interaction; public health relevance; research study; response; sialosyl-T antigen; tumor; tumor progression

DESCRIPTION (provided by applicant): This is a competing continuation of a RO1 project "IRS-1 JCV T-antigen interaction in Cerebellar Tumors". The results from previous founding period demonstrate that Insulin Receptor Substrate 1 (IRS-1), which is the major cytosolic component of the IGF-I receptor signaling system, can translocate to the nucleus. Importantly, we have detected nuclear IRS-1 in medulloblastoma cell lines and in medulloblastoma clinical samples, which were positive for polyomavirus JC large T-antigen. Our further studies demonstrate that nuclear IRS-1 can inhibit faithful component of DNA repair (homologous recombination DNA repair, HRR) via a direct interaction between IRS-1 and Rad51 detected at the sites of damaged DNA. We also demonstrated that the interference with DNA repair by nuclear IRS-1, and subsequent accumulation of mutations may happen in the absence of JCV T-antigen. In this respect, estrogen receptor beta (ER¿), which is highly expressed in medulloblastomas, can translocate IRS-1 to the nucleus in JCV T-antigen negative cells. These findings indicate that the interaction between IRS-1 and ER¿ could be engaged in cellular responses to DNA damage. It also suggests that inhibition of the faithful DNA repair by nuclear IRS-1 is a common event, not restricted to the medulloblastoma cases which are JCV positive. Therefore, we have developed a research plan which will analyze fundamental mechanisms leading to the development of genomic instability in medulloblastomas. The proposal is founded on the hypothesis that highly active ER¿ found in medulloblastoma mediates translocation of IRS-1 to the nucleus causing inhibition of HRR, unfaithful DNA repair, and the development of genomic instability in actively growing medulloblastoma cells. To test this hypothesis, we have developed a series of experiments, which are outlined in three Specific Aims. In Aim #1, we will evaluate ER¿ effects on IGF-I dependent and IGF-I -independent components of HRR, and will characterize binding and subcellular localizations among ER¿, IRS-1 and Rad51 in human medulloblastoma cell lines. In Aim#2, we will evaluate how pharmacological and molecular manipulations with the IGF-IR-IRS-1-E¿ signaling axis affect DNA repair fidelity in vitro and tumor progression in RCAS/tv-a transgenic mice. Finally, in Aim #3, we will analyze protein levels, subcellular localization and co-localization among IRS-1, ER¿, and Rad51 in clinical samples obtained from patients with medulloblastoma. Since DNA damaging agents are frequently used to eliminate CNS tumors including medulloblastoma, defects in DNA repair fidelity may have strong mutagenic consequences. This in turn may result in tumor progression or tumor recurrences after genotoxic treatment. Better understanding of the signaling cross-talk between IRS-1, ER¿ and DNA repair proteins is expected to provide new molecular targets for future therapeutic strategies against genomic instability, which is common in medulloblastoma patients. PUBLIC HEALTH RELEVANCE Medulloblastomas are cerebellar tumors of the childhood, in which the spread of tumor cells within central nervous system (CNS) is associated with poor prognosis. Since DNA damaging agents are frequently used to eliminate medulloblastoma, defects in DNA repair mechanisms are strongly suspected to cause accumulation of mutations contributing to tumor progression or tumor recurrences. The proposed studies of DNA repair mechanisms in association with cell signaling pathways are critically important for the development of new therapeutic strategies against malignant dissemination of these cerebellar tumors in CNS.

描述（由申请人提供）：这是 RO1 项目“IRS-1 JCV T 抗原在小脑肿瘤中的相互作用”的竞争性延续。先前创建时期的结果表明胰岛素受体底物 1 (IRS-1)，即胰岛素受体底物 1。 IGF-I 受体信号系统的主要胞质成分可以转位至细胞核，重要的是，我们在髓母细胞瘤细胞中检测到了核 IRS-1。我们的进一步研究表明，核IRS-1可以通过IRS-1之间的直接相互作用抑制DNA修复的忠实成分（同源重组DNA修复，HRR）。我们还证明，在缺乏 JCV T 抗原的情况下，核 IRS-1 会干扰 DNA 修复，并导致随后的突变积累。受体β (ER¿) 在髓母细胞瘤中高度表达，可以将 IRS-1 易位到 JCV T 抗原阴性细胞的细胞核中。这些发现表明 IRS-1 和 ER¿ 之间的相互作用。这也表明核 IRS-1 抑制忠实的 DNA 修复是一种常见事件，并不限于 JCV 阳性的髓母细胞瘤病例。将分析导致髓母细胞瘤基因组不稳定性发展的基本机制。该提案基于高度活跃的 ER¿在髓母细胞瘤中发现的 IRS-1 介导 IRS-1 易位到细胞核，从而抑制 HRR、不忠实的 DNA 修复以及活跃生长的髓母细胞瘤细胞中基因组不稳定性的发展。三个具体目标 在目标 #1 中，我们将评估 ER¿对 HRR 的 IGF-I 依赖性和 IGF-I 独立成分的影响，并将表征 ER¿ 之间的结合和亚细胞定位、IRS-1 和 Rad51 在人髓母细胞瘤细胞系中的作用 在 Aim#2 中，我们将评估如何使用 IGF-IR-IRS-1-E¿最后，在目标 #3 中，我们将分析 IRS-1、ER¿ 之间的蛋白质水平、亚细胞定位和共定位。和Rad51在临床上从髓母细胞瘤患者获得的样本中，由于消除DNA损伤剂经常用于治疗包括髓母细胞瘤在内的中枢神经系统肿瘤，DNA修复保真度的缺陷可能会产生强烈的诱变后果，这反过来会导致基因毒性治疗后的肿瘤进展或肿瘤复发。更好地理解 IRS-1、ER¿ 之间的信号串扰。 DNA 修复蛋白有望为未来针对基因组不稳定性的治疗策略提供新的分子靶点，这在公共卫生相关性髓母细胞瘤是儿童小脑肿瘤，其中肿瘤细胞在中枢神经系统 (CNS) 内扩散。由于 DNA 损伤剂经常用于消除髓母细胞瘤，因此强烈怀疑 DNA 修复机制的缺陷会导致导致肿瘤进展或肿瘤的突变积累。所提出的与细胞信号通路相关的 DNA 修复机制的研究对于开发针对中枢神经系统小脑肿瘤恶性播散的新治疗策略至关重要。